Integrins are a large family of homologous transmembrane linker proteins, and are the principal receptor proteins on animal cells which bind extracellular matrix proteins such as collagen, fibronectin, and laminin. Integrins consist of two noncovalently associated transmembrane glycoprotein subunits, called .alpha. and .beta., both of which contribute to the binding of the target extracellular matrix protein. The binding of integrins to their ligands requires the presence of extracellular divalent cations (Ca.sup.2+ or Mg.sub.2+, depending on the integrin); multiple divalent-cation-binding domains are present in the large extracellular part of the .alpha. chain. Integrin receptors undergo reversible activation due to ligand binding or cellular stimulation; activation results in conformational changes in the integrin extracellular domains, reorganization of intracytoplasmic connections, and redistribution of integrins on the cell surface. (See Diamond and Springer, Curr. Biol. 4:506 (1994); Li et al., J. Cell Biol. 129:1143 (1995); Yednock et al., j. Biol. Chem. 270:28740 (1995)). About 20 integrin heterodimers, made from 9 types of .beta. subunits and 14 types of .alpha. subunits have been defined.
As noted above, binding of integrins to their ligands depends on extracellular divalent cations. This property can be used to purify integrins, for example, by passing detergent-solubilized plasma membrane proteins over an affinity column containing an extra-cellular matrix protein, and then eluting the bound integrins from the column by washing in a divalent-cation-free solution. The Molecular Biology of the Cell, 3d ed., Alberts et al. (eds.), Garland Publishing Inc., New York, 1994, p. 996-1000.
.beta.3 integrins are found on a variety of cells and are known to bind several matrix proteins, including fibrinogen. Blood platelets contain the .alpha.IIb.beta.3 fibrinogen receptor (also known as GPIIb IIIa). Platelet integrins can be activated by contact with a damaged blood vessel or by any of a number of soluble signaling molecules. Activation alters the conformation of the extracellular domain of .beta.3 integrins, so that the extracellular domain becomes able to bind fibrinogen with high-affinity, promoting platelet aggregation and blood clot formation. In unstimulated platelets the majority of .alpha.IIb.beta.3 is present in an inactive conformation or low affinity state, and is unable to bind soluble ligands. Platelet agonists through their respective receptors transduce a cascade of intracellular signals ultimately leading to the conversion of .alpha.IIb.beta.3 into an active, high affinity state that is capable of binding soluble ligands.
Shattil et al. J. Cell Biol. 131:807 (1995), using the yeast two-hybrid system, recently identified a novel protein termed .beta.3-endonexin that interacts with the cytoplasmic domain of the .beta.3 integrin.
U.S. Pat. No. 5,523,209 to Ginsberg and O'Toole (Jun. 4, 1996) describes a method using cell culture assays to screen compounds to identify inhibitors of integrin activation.